Nitrates of bis-platinum complexes with polyamine ligands

ABSTRACT

Nitrates of Bis-platinum complexes with polyamine ligands of formula (I) having antitumor activity and pharmaceutical compositions containing said compounds as active principles.

CROSS REFERENCE TO RELATED APPLICATION

This application is a 35 U.S.C. 371 National Phase Entry Applicationfrom PCT/EP00/06288, filed Jul. 5, 2000, and designating the U.S.

The present invention relates to compounds of formula (I),

in which n is zero or the integer 1.

Furthermore, the present invention relates to the pharmaceuticalcompositions containing said compounds as active principles.

The compounds of formula (I) are bis-platinum complexes with polyamineligands having antitumor activity against both resistant andnon-resistant cells to cisplatin.

TECHNOLOGICAL BACKGROUND

WO 98/03519 discloses bis-platinum complexes with polyamine ligands(spermine and spermidine) which are the same as those of formula (I),but are salified with chloride anions instead of nitrate ones.

Said compounds, respectively represented by the formula:

have potent cytotoxic activity against cis-platinum resistant tumourcells, such as the L1210/CDDP and A2780/CDDP cell lines. The in vitroactivity was further confirmed in vivo, in murine tumors experimentalmodels.

The disclosed compounds have not, however, ideal solubility, stabilityand purity characteristics in view of the envisaged pharmaceutical use.

DISCLOSURE OF THE INVENTION

It has now been found that the compounds of formula (I), which are thenitrates of the above mentioned compounds A and B, have improvedsolubility, stability and purity characteristics, so that they areparticularly suitable to the formulation of pharmaceutical compositions,above all for the parenteral use.

The compound of the invention of formula I in which n is 1 (Compound 1,spermine derivative) and the compound of formula I in which n is zero(Compound 2, spermidine derivative), can be prepared starting from thecorresponding A and B chloride salts, respectively, by reaction withalkaline nitrates or silver nitrate in aqueous solutions.

Sodium nitrate is preferably used in high concentrations, typicallyranging from 2 to 5 M, in the presence of strong inorganic acids such asHCl, in concentrations ranging from about 1 to about 10 mM. The nitratesalts precipitate in the form of crystalline solids which can be washedwith alcoholic or water-alcoholic solutions. The preparation of thestarting Compound A and Compound B chloride salts is reported inexamples 16 and 14, respectively, of WO 98/03519 which is hereinincorporated for reference.

The compounds of formula (I), when administered to humans and animalsbearing tumors which can be treated with cis-platin or to which they areresistant, at doses ranging from 0.1 mg to 1.2 g per square metre bodyarea, are capable of inducing the regression of said tumors.

More generally, the compounds of the invention can be used for thetreatment of the same pathological conditions for which cis-platin isused. This includes the treatment of tumors, sensitization orenhancement of radiations [Douple et al., Cis-platin Current Status andDevelopments, Ed. A. W. Prestayk et al., Academic Press, 125 (1980);Douple et al., Platinum Metals Res., 29; 118 (1985)] and the treatmentof parasitic diseases such as African sleeping sickness [Farrell et al.,Biochem. Pharmacol., 33, 961 (1984)].

Therefore, another object of the present invention consists ofpharmaceutical compositions containing a therapeutically effectiveamount of at least one compound of formula (I) in mixture withconventional carriers and excipients.

The effective dosage of the compounds of the invention can be determinedby expert clinicians according to conventional methods. The relationshipbetween the dosages used for animals of various species and sizes andthose for humans (on the basis of mg/m² body area) is described byFreirech et al., Quantitative Comparison of Toxicity of AnticancerAgents in Mouse, Rat, Hamster, Dog, Monkey and Man, Cancer Chemother.Rep., 50, N. 4, 219-244 (1986).

Usually, however, the patient will receive doses from 0.1 to 1200 mg/kgbody weight of the complex, with a dosage regimen which will varydepending on various factors which are well known to the expertclinicians.

The treatment regimen can suitably be varied, as it is well known to theexpert clinician, according to the type of tumor to be treated and theconditions of the patient.

The compounds of the invention will be administered through theparenteral or oral routes.

The pharmaceutical compositions for the parenteral administrationcomprise sterile saline solutions, as defined above, or sterile powdersfor the extemporary preparation of the solutions, as well as oilypreparations for intramuscular (im) or intraperitoneal (ip)administrations.

The compounds of the invention are preferably administered as sterileaqueous solutions, optionally containing sodium chloride in suitableconcentration (0.1-0.9 mg/ml). The solutions are preferably administeredby the intravenous (iv) or intra-arterial (ia) routes, even though otheradministration forms can be used in particular cases.

Useful pharmaceutical compositions for the oral administration can besyrups or similar liquid forms, as well as solid forms such as tablets,capsules and the like.

The pharmaceutical compositions according to the present invention areprepared following known methods, such as those reported in Remington'sPharmaceutical Sciences Handbook, XVII Ed., Mack Pub., N.Y., U.S.A.

Sometimes it can prove advantageous to administer the platinum complexesof the present invention together with one or more agents which enhancethe antitumor activity or relieve the undesirable side effects which maybe associated with the platinum complex therapy. For example, theplatinum complexes of the present invention can be administered togetherwith reduced glutathione, as disclosed in GB 2,174,905 and in U.S. Pat.No. 4,871,528.

Furthermore, it can be advantageous to administer the platinum complexesof the present invention in combination with other platinum complexeshaving antitumor activity.

Therefore, a further object of the present invention consists ofpharmaceutical compositions containing at least one compound of formula(I) in combination with a platinum complex having antitumor activity.

A further object of the present invention consists of the use of thecompounds of formula (I) for the preparation of pharmaceuticalcompositions for the treatment of mammals bearing tumors which can betreated with cis-platinum or resistant to cis-platinum.

THE INVENTION IS FURTHER ILLUSTRATED BY THE FOLLOWING EXAMPLES. EXAMPLE1 Preparation of bis{trans(diammine)(chloro) Platinum(II)}-μ-(1,8-diamino-4-azaoctane-N¹,N⁸)dinitrate salt.HNO₃, (Compound 2)

A solution of bis{trans(diammine)(chloro)platinum(II)}-μ-(1,8-diamino-4-azaoctane-N¹, N⁸)dichloride salt.HCl prepared according to example 14 of WO 98/03519,(0.306 g, 0.391 mmol) in 8 mM HCl (15 ml) at room temperature is addedwith active charcoal (Norit A, 50 mg) and the mixture is stirred for 30minutes. Charcoal is filtered off and the resulting clear filtrate iscooled to 10° C. A 4N NaNO₃ solution (1.5 mL, 5.87 mmol) is added andwithin a few minutes a white solid precipitates. The mixture is stirredfor 3 hours, then the precipitated solid is filtered, washed withH₂O/MeOH 1:1 (2×3 mL), with MeOH (3 mL) and dried under vacuum (35° C.,2 h) to obtain 0.25 g (74% yield) of bis{trans(diammine)(chloro)platinum(II)}-μ-(1,8-diamino-4-azaoctane-N¹, N⁸) dinitratesalt.HNO₃, (Compound 2).

Analytical Data

¹H-NMR (δ; D₂O): 1.75 (bt, 4H); 2.15 (m, 2H); 2.75 (m, 4H); 3.10 (bt,4H); HPLC: rt (Compound 2)=6 min (97.2 area%); Elemental analysis:calculated %: C, 9.76; H, 3.74; N, 16.26; Cl 8.23; Pt 45.29. found %: C,9.84; H, 3.76; N, 15.67; Cl 9.04; Pt 45.15.

EXAMPLE 2 Preparation of bis{Trans (diammine)(Chloro)platinum(II)}-μ-(1,12-diamino-4,9-diazadodecane-N¹,N¹²)dinitrate Salt.2HNO₃, (Compound 1)

A solution ofbis{trans(diammine)(chloro)platinum(II)}-μ-(1,12-diamino-4,9-diazadodecane-N¹,N¹²) dichloride salt.2HCl, prepared according to Example 16 of WO98/03519 (0.71 g, 0.811 mmol) in 8mM HCl (60ml) at room temperature isadded with active charcoal (Norit A, 50 mg) and the mixture is stirredfor 30 minutes. Charcoal is filtered off and the resulting clearfiltrate is cooled to 10° C. A solution of 4N NaNO₃ (3 mL, 12 mmol) isadded and within a few minutes a white solid precipitates. The mixtureis stirred for 13 hours, then the precipitated solid is filtered, washedwith H₂O/MeOH 1:1 (2×3 mL), with MeOH (3 mL) and dried under vacuum (35°C., 2h) to obtain 0.6 g (75% yield) ofbis{trans(diammine)(chloro)platinum(II)}-μ-(1,12-diamino-4,9-diazadodecane-N¹,N¹²) di-nitrate salt.2HNO₃, (Compound 1).

Analytical Data

¹H-NMR (δ; D₂O): 1.75 (m, 4H); 2.10 (bq, 4H); 2.80 (bq.4H); 3.15 (bt,8H); HPLC: rt (Compound 1)=11 min (98.8 area%); Elemental analysis:calculated %: C, 12.24; H, 4.11; N, 17.12; Cl 7.22; Pt 39.65. found %:C, 12.43; H, 4.09; N, 17.02; Cl 7.26; Pt 39.71.

EXAMPLE 3

FIG. 1a shows the chromatographic profile obtained withbis{trans(diammine)(chloro)platinum(II))}-μ-(1,8-diamino-4-azaoctane-N¹,N⁸) dichloride salt.HCl, (Compound B, prepared as described in example14 of WO 98/03519) compared with the chromatographic profile of figure1b obtained with the corresponding bis{trans(diammine)(chloro)platinum(II)}-μ-(1,8-diamino-4-azaoctane-N¹, N⁸) dinitratesalt.HNO₃ (Compound 2) of example 1, using the following chromatographicsystem:

column: Lichrospher RP8 (5 μm), 125×4 mm

eluent A: H₂O/CH₃CN 80/20 sodium octanesulfonate (2.5 g/L) pH 2.7 with85% H₃PO₄ eluent B: H₂O/CH₃CN 45/55 sodium octanesulfonate (2.5 g/L) pH2.7 with H₃PO₄ 85% Gradient: eluent A/eluent B 50/50 0-24′) to eluentA/eluent B 25/75 (26′-50′) flow: 1 mL/min detection UV (225 nm)

EXAMPLE 4

FIG. 2a shows the chromatographic profile obtained withbis{trans(diammine)(chloro)platinum(II)}-μ-(1,12-diamino-4,9-diazadodecane-N¹,N¹²)dichloride salt.2HCl, (Compound A, prepared as described in example16 of WO 98/03519) compared with the chromatographic profile of FIG. 2bobtained with the correspondingbis{trans(diammine)(chloro)platinum(II)}-μ-(1,12-diamino-4,9-diazadodecane-N¹,N¹²) dinitrate salt.2HNO₃ (Compound 1), of example 2, using thechromatographic system described in example 3.

EXAMPLE 5

The following table reports the HPLC data concerning the stability ofCompound 1 and of Compound A (prepared as described in example 16 of WO98/03519) following heating at 40° C. for 12 days under atmosphericpressure.

The used chromatographic method is that described in Example 3 andExample 4. The values of retention times are in percent area.

Compound 1 Compound A Retention 12 days 12 days times (min) t = 0 40° C.t = 0 40° C. MAIN IMPURITIES mean peak 98.5 98.4 92.5 89.8  7 0.1 0.10.1 0.8  8 0.1 0.1 0.2 1.6 12 0.1 0.2 0.3 0.6 14 0.8 0.9 1.3 0.8 30 <0.1<0.1 2.3 2.8 31 n.d.* 0.2 0.7 1.7 *n.d. = not detectable

What is claimed is:
 1. A compound of formula (I),

in which n is zero or the integer
 1. 2. A process for the preparation ofa compound of claim 1, comprising reacting the corresponding chloridesin solutions in strong organic acids with an aqueous solution of analkaline or silver nitrate.
 3. A process as claimed in claim 2, in whichthe inorganic acid is hydrochloric acid in 1 to 10 mM concentration. 4.A process as claimed in claim 2, in which the alkaline nitrate is sodiumnitrate.
 5. A Pharmaceutical composition containing as active ingredienta therapeutically effective amount of at least one compound of claim 1,together with conventional carriers and excipients.
 6. A method oftreating in a patient a tumor treatable with or resistant tocis-platinum, comprising administering to the patient an anti-tumoreffective amount of a compound of claim 1.